Association Of The Microsatellite Alterations On Human Chromosome 3p, 9p And 14q With Renal Cell Carcinoma

【Objective】To explore the relation between microsatellite alterations on human chromosome 3p, 9p and 14q in renal cell carcinoma and clinicopathological characteristics,and to find the prognostic biomarkers in clear cell renal cell carcinoma.【Methods】1.82 RCC specimens were obtained from 69 RCC patients.The representative markers from D14S1426,D14S617,D14S260,D3S1289,D3S1560,D3S1566,D3S1300, D3S1633,D9S171,D9S1749,D9S157 and D9S168 were selected to investigate microsatellite alterations such as MSI and LOH by the best method which was selected from PCR-PAGE-EB staining,PCR-denatured PAGE-silver staining,PCR-SSCP-EB staining,and sequencing.The relation between microsatellite alterations and clinicopathological characteristics were analyzed by x~2 test.2.48 patients with primary CCRCC surgically treated by nephrectomy were followed up by telephone.The initial event was the first diagnosis of CCRCC.The end point was metastasis or death related to CCRCC.The visiting end date was April 23th,2007.Survival curves were constructed using the Kaplan-Meier method and differences were assessed by the log rank test.Cox proportional hazards regression models were used to examine the relative impact of the multi-variables by forward stepwise likelihood ratio to further reduce the number of variables in the Cox model.For assessing and comparing the Cox model a likelihood test with a significance level of 0.05 was used for inclusion and 0.10 for exclusion of variables.【Results】1.The result was the same by the above four detecting methods in the same patient. Comparing PCR-PAGE-EB staining with sequencing in 27 pairs of tissues,I found Kappa=0.706 and agreement of the two methods was good.The five representative markers—D14S1426,D14S617,D3S1289,D3S1560 and D9S168 were selected on base of PCR success rate and microsatellite alteration rate in 31 patients' tissues.I found LOH in D3S1289 when one RCC patient had RCC again on the right kidney was different from the first time of diagnosis on the left kidney 8 years ago.No stastically significant difference was showed between 12 patients' primary RCC tissues and the matched metastatic tissues (P＞0.05).The frequency of MSI in D14S617,D14S1426,D3S1289,D3S1560 and D9S168 in 69 RCC patients were respectively 5.9%(4/68),4.5%(3/67),1.6%(1/63), 8.2%(5/61),19.7%(13/66);the frequency of LOH were respectively 10.3%(7/68), 3.0%(2/67),11.1%(7/63),9.8%(6/61),4.5%(3/66);all the frequency of alterations were respectively 16.2%(11/68),7.5%(5/67),12.7%(8/63),18.0%(11/61),24.2%(16/66).The frequency of more than two markers'alterations was 21.7%(15/69).Sex,age,pathology, TNM staging,and metastatic status were not associated with MSI and LOH in D14S617, D14S1426,D3S1289,D3S1560 and D9S168(P＞0.05).2.The Kaplan-Meier analysis indicated that development of CCRCC metastasis was significant dependent on the alteration of D9S168(P=0.001) and not significant dependent on the alterations of D14S617(P=0.207),D14S1426(P=0.331),D3S1289(P=0.918),and D3S1560(P=0.981) by log-rank test.The Kaplan-Meier analysis indicated that overall survival of CCRCC was significant dependent on the alteration of D9S168(P=0.003) and not significant dependent on the alterations of D14S617(P=0.237),D14S1426(P=0.218), D3S1289(P=0.965),and D3S1560(P=0.974) by log-rank test.Multivariate analysis by Cox proportional hazard model revealed D9S168 was an independent prognostic factor for CCRCC metastasis and CCRCC-related death.Hard ratios for patients with D9S168 alteration were 15.08(95%CI,1.67～136.15) for metastasis,and 13.37(95%CI,1.49～120.16) for tumor-related death.【Conclusion】1.PCR-PAGE-EB staining is a simple,cheap,and reliable method to detected microsatellite alterations.2.The development of RCC is controlled by many risks.It is possible that there are some anti-oncogene or mismatch repair gene related RCC near D14S617,D14S1426, D3S1289,D3S1560 and D9S168.3.It is possible that metastasis is positive for RCC and the microsaellite alterations just impact on the speed.D9S168 was an independent prognostic factor for CCRCC metastasis and CCRCC-related death.The tumor cells which have the alteration of D9S168 have more highly invasive capability than other cells which don't.Maybe some gene related CCRCC metastasis near D9S168 is usefull for gene therapy of RCC.